Impact of CircRNAs on Ischemic Stroke

Circular RNA (circRNA) is a recently discovered class of endogenous non-coding RNA that is generated by cyclization, driven by intron pairing, and regulated by related regulators. An important biological function of CircRNA is acting as a molecular sponge to significantly alter miRNA levels over a short period. Several studies have shown that circRNA is closely related to stroke. Therefore, a better understanding of CircRNA function and regulatory mechanism in ischemic brain will help us for the early detection, early diagnosis, and early treatment of stroke. Here, we summary the biological characteristics, expression of circRNA, and its effect on outcome after ischemic stroke.


Characteristics of circRNA
CircRNAs, a special class of endogenous non-coding RNA, are usually in vitro and in vivo produced by exon or intron cyclization splicing or reverse splicing. According to the genomic origin, CircRNA can be divided into three categories: annular intron RNA (ciRNA), exon RNA (eCircRNA), and exon-intron CircRNA (eiCircRNA). Notably, based on the positional relationship between circRNA and adjacent coding RNA, the aforementioned three CircRNAs can be further divided into five subtypes: exon, intron, antisense, justice overlap, and intergenic [5].
Unlike the structure of linear RNA or other types of RNA, circRNA has a closed covalent ring structure endowing the verified circRNA with several characteristics [6]. CircRNA is less sensitive to exonuclease and more stable than linear RNA because of the structural advantage of lacking free 3' and 5' ends. Human circRNAs were initially found in the 1990s. At the time, they were considered aberrant splicing products generated by splicing mistakes. Furthermore, there is generally a low amount of circRNA, and standard methods for investigating linear RNA are ineffective for researching circRNAs. Therefore, their function is understudied [7]. CircRNA was previously found in various species, thanks to advancements in biochemical enrichment approaches and total transcriptome sequencing techniques (RNA-seq).

A single circRNA acts as a miRNA sponge
Theoretically, circRNA binds with miRNA competitively like a sponge through base complementarity, which is called "miRNA sponge action" [8]. The connections between circRNA and the binding site of miRNA can affect biological function. It has been found that circRNA-CDR1, ciRS-7, circ-ITCH, circ-SRY can act as miRNA sponges [9]. For example, the mammalian sexdetermining gene murine SRY (sex-determining region Y) can encode testicular specific circular transcripts. Cirs-7 is a typical circRNA containing more than 70 miR-7 binding sites, which exists in many tissues, especially brain tissues [9].

Interaction with RNA binding protein
The most famous protein that interacts with RNA molecules is RNA binding protein (RBP). CircRNA competitively binds to the substrate-binding site of RBP through the mode of storing and transporting RBP and then regulates RBP activity [10]. CircRNA can operate as a miRNA sponge to suppress miRNA activity, engage in target gene splicing, translate genes into proteins, and interact with RBP. RBP interacts with circRNA and stimulate CircRNA production, while circRNA can control the functional activity of RBP. CircRNA can act as a protein sponge or bait to affect cell function, regulate gene transcription, inhibit cell cycle processes, induce apoptosis and promote cell proliferation and survival [11].

Regulating the expression of proteins and genes
As we mentioned earlier, one kind of regulatory circRNA, called exon-intron circRNA (EIciRNA), plays a role in transcriptional regulation. EiCircRNA is a multi-exon circRNA that contains one or more non-spliced inserted introns. Some eiCircRNAs located in the nucleus can promote the transcription of their parent genes by interacting with the spliceosome component U1 small ribonucleoprotein (snRNP) [12]. Previous study has demonstrated that EIciRNA can inhibit parent gene transcription by interacting with host U1 snRNP and RNA Pol II to compete with mRNA and influence protein translation [10]. According to Abdelmohsen et al., circRNA (circPABPN1) and its corresponding mRNA competed in the RBP HuR, affecting protein production [13]. Another study [14] found that circANRIL inhibited rRNA processing and ribosomal function while activating p53 in human vascular endothelial cells and macrophages 60s via binding to the ribosomal assembly factor pescadillo homolog 1 (PES1). CircRNAs regulate ribosomes and play a certain role in protein expression. These studies all confirmed that ribosomes regulated by CircRNAs play a certain role in protein expression.

Other functions
It is also important to point out that there are many other potential features of circRNA, such as competing with mRNA clipping (e.g., the splicing factor muscle blind, mbl) [10]. In addition, it has been reported that circRNA can also affect cell differentiation, participate in pseudogene formation, the regulation of intercellular signal pathway, and stress response, and consequently further impact the occurrence and development of stroke.

CircRNA expression profile after IS
Biomarkers are highly important for early detection and follow-up monitoring of IS, and they contribute to a thorough understanding of the disease's etiology and pathophysiology. MiRNAs and lncRNAs have been proposed as possible stroke biomarkers [15,16]. Furthermore, circRNAs have several functions in the onset and progression of IS and are gaining interest as a possible biomarker. Understanding the differential expression of circRNAs as the disease progresses will provide us new biomarkers for the diagnosis and prognosis of IS. Many studies have shown that the expression of circRNAs in brain and plasma is affected during the occurrence of cerebral infarction and the differential expression of circRNA in cells of oxygen glucose deprivation/re-oxygenation (OGD/R), an in vitro model of IS. Moreover, there are differences in the expression of circRNA in patients with different stroke etiology and in the non-ischemic area of IS.

Differential expression of circRNA in brain tissues after IS
Mehta et al. [17] developed the middle cerebral artery occlusion (MCAO) model in male C57BL/6J mice for the first time. At 6, 12, and 24 h after reperfusion, 1320 differential circRNAs were discovered, of which 283 cases changed at different reperfusion time courses. The qRT-PCR analysis revealed that circ-008018, circ-015350, and circ-016128 were upregulated, whereas circ -11137, circ-001729, and circ-006696 circRNAs were downregulated. Their primary biological and molecular activities include biological regulation, metabolic processes, cellular communication, and proteins, ions, and nucleic acids binding. Liu et al. [18] analyzed the mouse brain tissue microarray after MCAO for 45 min and found that 1027 circRNAs changed significantly in the brain tissue 48 h after reperfusion. By analysis of qRT-PCR, mmu-circRNA-40001, mmu-circRNA-013120, and mmu-circRNA-25329 are differential expressed. It regulates cell proliferation and survival mainly by participating in the regulation of the Rap1 pathway and Hippo pathway. Further analysis shows that these circRNA target genes may be involved in regulating different biological processes. Finally, the interaction network of circRNA-miRNA target genes includes 13 miRNAs and their corresponding genes.
Taken together, the changes of circRNAs were related to the genes associated with brain injury and recovery.

Expression profile of circRNA in the cerebral infarction area
Lu et al. [19] investigated circRNA expression in the blood of transient MCAO model in mice and confirmed the chosen circRNA in patients with acute IS (AIS). The findings revealed that 128, 198, 789 circRNAs were substantially altered 5 min, 3 h, and 24 h after IS, with their targeting genes linked to the Hippo signal pathway, extracellular matrix-receptor interaction, and fatty acid metabolism, respectively. Finally, circBBS2 and circPHKA2 were shown to be differently expressed in the blood of patients with AIS. HTS was used to compare the expression of circRNAs in peripheral blood mononuclear cells (PBMC) from five patients with AIS and five healthy subjects [20]. Compared with the normal control group, 521 circRNAs were differentially expressed, of which 373 circRNAs were upregulated, and 148 circRNAs were downregulated. QRT-PCR confirmed that eight circRNAs contained multiple microRNA binding sites. GO and KEGG analysis showed that abnormal expression of circRNAs was involved in many pathophysiological processes of AIS, especially inflammatory and immune processes. The connection between circRNAs and IS induced by MCAO was addressed by Duan et al. [21]. The findings revealed that 87 of the differentially expressed circRNAs had more than two-fold changes. circRNA. 17737, CircRNA.8828, and CircRNA.14479 were all substantially upregulated, whereas circRNA.1059, circRNA.9967, and circRNA.6952 were all significantly downregulated.
Li et al. [22] analyzed the circRNA expression patterns in three patients with AIS and three normal healthy controls in the Han population in southern China. They found that 2270 circRNAs were expressed differently, of which 659 were upregulated, and 1611 were downregulated. De circRNAs may participate in the pathophysiology of AIS through endocytosis, energy metabolism, apoptosis, the FOXO signaling pathway, platelet activation, neurotrophic factor signaling pathway, and VEGF signaling system. The qRT-PCR results showed that hsa-Circ-0005548 was significantly upregulated, while hsa-circ-0000607 and hsa-circ-0002465 were significantly downregulated. In addition, the area under the curve (AUC) values suggests that hsacirc-0000607 and hsa-circ-0002465 could be potential biomarkers of AIS, and hsa-Circ-0000607 may play a major role in the occurrence and development of AIS by regulating the mir-337-3p/BCL2 axis. Li et al. [23] explored circRNA expression profiles in five patients with LAA stroke and four controls. They found that 182 circRNAs were elevated and 176 were downregulated in patients with LAA stroke. qRT-PCR verified six differentially expressed circRNAs. These circRNAs are mainly involved in chromatin remodeling, autophagy, platelet activation, and brain precursor cell proliferation. The expression level of hsa-circRNA 0001599 was positively correlated with the National Institutes of Health Stroke Scale score (NIHSS) and infarct volume. The area under the receiver operating characteristic curve was 0.805, and the diagnostic sensitivity and specificity were 64.41% and 89.93%, respectively. There are differences in the expression of some circRNAs in the blood of patients with AIS, which may be useful as a diagnostic biomarker or potential therapeutic target of AIS.

CircRNA is differentially expressed in HT22 cells after OGD/R
Lin et al. [24] discovered that 15 circRNAs altered substantially, with 3 upregulated and 12 downregulated. MMU-circRNA-015947 expression is increased and can interact with microRNA to increase the expression of target genes. It contributes to the progression of ischemiareperfusion damage by involving in apoptosis, metabolism, and immune-related pathways. This research demonstrates that MMU-circRNA-015947 expression is implicated in the process of cerebral ischemia-reperfusion damage and possible for a new molecular target for therapeutic therapy.

The differencial expression of circRNA in stroke patients with different etiology
Aiora et al. [3] reported 219 differentially expressed circRNAs in patients with atherosclerotic and thrombotic stroke, of which 60 upregulated circRNAs showed more than quadruple expression. When comparing atherosclerosis and undetermined stroke, there were 226 circRNAs differentially expressed, 87 circRNAs upregulated, and 139 circRNAs downregulated, of which only one circRNA expression was more than quadrupled. When comparing thrombotic stroke with undetermined stroke, only 8 circRNAs were upregulated, and 9 circRNAs were downregulated. Differential expression of circRNAs in atherosclerosis and cardiac embolism was verified by qRT-PCR. It was found that only ubiquitin Amur52 ribosomal protein fusion product 1 (UBA52) gene HSA_circRNA_102488, which originated on chromosome 19, had statistically significant changes between different etiological subtypes, and the RBP site of hsa_circRNA_102488 was clustered around AGO2 and FUS protein. Finally, functional analysis showed that differentially expressed circRNAs mainly interacted with stroke-related miRNAs.

Differential expression of circRNA in the nonischemic area after IS
According to Li et al. [25], a total of 2659 circRNAs altered substantially in the ipsilateral thalamus of adult male C57BL/6J mice in the persistent distal MCAO compared to the control group. Among these, 73 circRNAs altered substantially after stroke. CircRNAs are shown to play key roles in subsequent thalamic degeneration and remodeling following a focal cortical acute infarction, according to GO and KEGG analyses. This is the first study to explore circRNA expression in the non-ischemic region of an IS, implying that circRNAs might be a therapeutic target for decreasing subsequent distal neurodegeneration following stroke.
In summary, these studies show that various circRNAs are differentially expressed after the onset of AIS, suggesting that circRNAs have potential application value in the diagnosis, treatment, and prediction of AIS (Table 1).

Role of circRNA in IS
CircRNA affects the occurrence and rehabilitation of IS in many ways. The role of circRNA in IS needs to be further elucidated and reviewed. Hence, in this section, the potential function and interaction of circRNAs after IS are summaried with a special focus on ischemic brain injury, protection of the blood-brain barrier, inhibition of apoptosis, neuroprotection, and neuroinflammation.

CircRNAs and atherosclerosis
Atherosclerosis is a chronic inflammatory disease of the vascular wall caused by many factors and is the main pathological cause of cardiovascular disease and stroke. In addition, the main causes of IS are carotid plaque rupture and thrombosis. Therefore, understanding the formation of carotid plaque may be helpful to predict and prevent cardiovascular and cerebrovascular events. We aimed to understand the role of circRNAs in serum exons of patients with stable plaque atherosclerosis (SA) and unstable/fragile plaque atherosclerosis (UA). Wen et al. [26] studied the effect of circRNA on the behavior of human umbilical vein endothelial cells (HUVECs) and the mechanism of plaque instability in AS. They found significant differences in the expression profiles of circRNA in serum exons of SA and UA, indicating that circRNA may play a role in carotid plaque instability. In patients with UA, the expression of circRNA-0006896 was positively correlated with triglycerides, low-density lipoprotein cholesterol (LDL-C), and C-reactive protein levels and negatively correlated with albumin levels. However, in patients with SA, the expression of CircRNA-0006896 was positively correlated with LDL-C. These findings suggest that the circRNA-0006896/ miR-1264/DNMT1 axis plays an important role in carotid plaque instability by regulating endothelial cell behavior.
In addition, circRNA-0006896 may be a therapeutic target for controlling JNK/STAT3 signal transduction in HUVEC. The phenotypic transformation of vascular smooth muscle cells (VSMC) is very important in the pathogenesis of atherosclerosis. Kang et al. [27] identified CDK6 as a key regulator of atherosclerosis, and CDK6 gene knockout inhibited the proliferation of HASMC and HUASMC cells. In addition, the circHIPK3/miR-637/CDK6 axis plays an important regulatory role in the proliferation and apoptosis of VSMC. We believe that circRNA is helpful to develop a simple and noninvasive early screening tool for vulnerable plaques to achieve early prediction and diagnosis of stroke. For example, serum Circ-284: miR-221 has been used as a biomarker for the diagnosis of carotid plaque rupture and stroke [28].

Ischemic brain injury
Using a circRNA microarray, it was discovered that the level of circHECTD1 was considerably elevated in ischemic brain tissue after transient MCAO in mice and plasma samples of AIS patients [29]. Mechanistically, circHECTD1 acts as an endogenous MIR142 (microRNA 142) sponge, inhibiting MIR142 activity and therefore inhibiting TIPARP (TCDD inducible poly [ADP-ribose] polymerase) production and astrocyte activation via macroautophagy/autophagy. Circhetd1 suppression decreased the infarct size, relieved brain injury, and improved astrocyte activation in transient MCAO animals. CircHECTD1 and its coupling mechanism could play a role in developing cerebral ischemia, giving substantial evidence to promote circHECTD1 as a novel stroke biomarker and therapeutic target. Wu and colleagues [30] reported an enhanced level of circTLK1 in the brain tissue of focal ischemia-reperfusion mice and in the plasma of patients with AIS. CircTLK1 ablation can reduce the infarct volume, minimize neuronal damage, and ameliorate neurological deficits. Furthermore, circTLK1 acted as an endogenous miR-335-3p sponge, inhibiting miR-335-3p activity and increasing 2,3,7,8tetrachlorodibenzo-p-dioxin-inducible poly (ADP-ribose) polymerase expression, leading to an aggravation of neuronal damage. These results further confirm that circTLK1 is directly associated with IS, indicating the critical role of circRNA on stroke prognosis. The retina is thought to be an outgrowth of brain tissue. However, it is currently unclear if circRNAs can be utilized as common regulators and diagnostic markers for brain and retinal neurodegeneration. Jiang et al. [31] discovered that silencing cGLIS3 reduced ischemia-induced retinal neurodegeneration and that cGLIS3 regulated neuronal cell damage by acting as a sponge for miR-203 and that its level was controlled by EIF4A3. From the standpoint of circRNA, this discovery provides molecular proof that the retina is a window into the brain. cGLIS3 is a common regulator as well as a diagnostic marker of cerebral and retinal neurodegeneration.

Protect the blood-brain barrier and inhibit apoptosis
Bai et al. [32] found that circRNA DLGAP4 acts as an endogenous microRNA-143 sponge to inhibit the activity of miR-143. The level of circDLGAP4 in the plasma of patients with AIS and the mouse stroke model significantly decreased. Moreover, in vivo experimental results showed that upregulating the expression of circDLGAP4 could significantly reduce the neurological deficit, infarct size, and blood-brain barrier injury. The in vitro results showed that over-expression of circDLGAP4 could reduce the downward trend of protein caused by oxygen OGD/R in mice and reduce the expression of mesenchymal cell markers to inhibit the transformation of endothelial cells into mesenchymal cells and reduce the damage of the blood-brain barrier. Furthermore, Zhao et al. [33] showed that the expression of circ-0072309 was significantly decreased while miR100 was significantly increased in the serum of patients with IS and the ischemic hemisphere of MCAO mice. Furthermore, they showed that miR-100 regulates cell survival and apoptosis by directly binding to mTOR, suggesting that he circ_0072309-miR-100-mTOR regulatory axis could be a potential target for the treatment of IS. More evidence has been found that circ_016719 can be directly targeted to miR-29c and control the expression and function of MAP2k6 associated with apoptosis [34]. As the overexpression of circCCDC9 can protect the blood-brain barrier and inhibit apoptosis via inhibiting the Notch1 signal pathway, circCCDC9 is considered another new potential therapeutic target for cerebrovascular protection in the acute phase of IS [35].

Neuroprotection
There is a significant drop in circMH1 levels in the plasma and periinfarct cortex of photothrombotic (PT) stroke mice and the plasma of patients with AIS. As circMH1 might promote neural plasticity while inhibiting glial cell activation and peripheral immune cell infiltration, circMH1 therapy may improve functional recovery in mice and monkeys following stroke. CircSCMH1 interacts with the transcription factor MeCP2, enabling the inhibition of MeCP2 target gene transcription. Based on such theoretical findings [36], Wang et al. [37] examined CircHIPK2's involvement in neural stem cell (NSC) differentiation. In vitro, suppressing CircHIPK2 aided NSCs in their differentiation to neurons but had no impact on astrocyte differentiation. After stroke onset, microinjected NSCs may migrate to the ischemic hemisphere in vivo. Si-circHIPK2NSCs improved neuronal plasticity in the ischemic brain, provided long term neuroprotection, and decreased functional impairments substantially. Dai et al. [38] discovered that circ-HECTD1 knockdown reduced TRAF3 expression via targeting miR-133b, reducing neuronal damage after cerebral ischemia. CircSHOC2 in ischemic preconditioning astrocyte exosome (IPAS-EXOs) effectively inhibits neuronal apoptosis and reduces neuronal damage with regulated autophagy on the miR-7670-3p/SIRT1 axis, which may develop IS treatment strategies [39]. A recent report discovered that hsaCirc0078299 and FXN might be considered as new biomarkers of IS to achieve neuroprotection and brain recovery from stroke [40].

Neuroinflammation, outcome and recurrence
For the neuroinflammation investigation, around 170 patients with AIS and 170 non-AIS controlled groups were selected to collect the PBMC and serum [41]. PBMC-Circ-DLGAP4 expression was negatively linked to PBMC-miR-143, NIHSS score, CRP, ESR, TNF-, IL-1, IL-6, IL-8, IL-17, and IL-22. It is also related to inflammation and miR-143 expression in patients with AIS. Circ-DLGAP4 has the potential to be used as a novel biomarker for AIS diagnosis and disease monitoring. Similarly, Peng et al. [42] investigated 160 patients with initial AIS and 160 non-AIS as controls and found that CircRNA HECTD1 expression was strongly associated with higher disease risk, disease severity, inflammation, and AIS recurrence. Xu Liu et al. [43] considered circ-STAT3 (signal transducer and activator of transcription) rs2293152 GG as an independent risk factor for stroke recovery. Subgroup analysis showed that the negative effects of rs2293152 GG genotype were higher in females, the elderly, and people with a history of hypertension. In addition, circRNA polymorphism was not associated with IS recurrence. The results show that circ-STAT3 may be a new biomarker to predict the functional outcome after stroke and is also an important factor in the recovery of IS. According to Chen et al., the expression of hsa-circ-0141720 in the serum of patients with ACI increased the most [44]. Further studies showed that the high expression of hsa-circ-0141720 was closely related to the NIHSS score, infarct volume, serum interleukin-6 (IL-6), and plasma C-reactive protein (hs-CRP). The high expression of hsa-circ-0141720 in the serum of patients with ACI is related to the severity of the disease, which can be used as a new serological index in the diagnosis of ACI.

Application of circRNAs in other aspects of IS
Identifying individuals at high risk of stroke-related infection (SAI) for preventive antibiotic treatment is essential for patients with AIS. Therefore, to determine whether circFUNDC1 may be a potential predictor of SAI, Zuo et al. [45] investigated 68 patients, of which 26 were infected and 42 were not. Compared to uninfected AIS patients, the level of circFUNDC1 in SAI patients was significantly elevated. The receiver characteristic (ROC) curve demonstrated the predictive significance of circFUNDC1. Furthermore, the level of circFUNDC1 and the number of neutrophils were shown to be positively correlated. In patients with SAI, the ratio of white blood cells to neutrophils was substantially greater than in non-SAI patients. Therefore, circFUNDC1 may be utilized to build a risk prediction model for SAI (Table 2) For the first time, Xiao et al. [46] demonstrated the comprehensive expression of exosomal circRNAs, showing their potential diagnostic and biological significance in LAA stroke. In peripheral exosomes, RNA-Seq revealed a total of 462 circRNAs, with 25 DE circRNAs. CircRNA competitive endogenous RNA (ceRNA) network and translatable analyses further showed the possible roles of exosomal circRNAs in LAA development. qRT-PCR confirmed two ceRNA pathways involving 5 circRNAs, 2 miRNAs, and 3 mRNAs. ROC curve analysis in the validation cohort indicated two circRNAs as potential new biomarkers, and a logistic model combining two and four circRNAs improved the AUC compared to the individual circRNAs.

CircRNAs after ischemia/reperfusion injury
Ischemia/reperfusion (I/R) damage occurs when blood perfusion returns after IS, causing the region of ischemic injury to extend and worsen. Brain tissue is extremely sensitive to I/R damage, and treatment of I/R damage as soon as possible is important to avoid severe consequences associated with neural cell death. CircRNAs have been shown to play a role in stroke and NSC modulation, and in the recovery of brain function. According to Yang B et al., in cerebral infarction, circTTC3 modulates CIR damage and NSCs via the miR-372-3p/TLR4 axis [47]. Furthermore, Zhang et al. [48] demonstrated that over-expression of hsa-Circ-camk4 increased cell death following OGD/R. Circ-camk4 works primarily via "sponging" miRNAs. The pathways most strongly involved in circ-camk4 regulation were those acting at glutamatergic synapses, as well as the MAPK signaling pathway, calcium signaling pathway, and apoptosis pathway, all of which are involved in neuronal cell response to I/R injury. These findings suggest that circ-camk4, as a type of circRNA, may play a key role in brain I/R injury.

Potential role of circRNAs in the treatment of IS
It is important to develop a new strategy for neuroprotection and neural tissue recovery after IS. Natural regulatory peptides can significantly affect brain activity and have high safety without side effects. Preparations generated from natural regulatory peptides have effectively created therapeutic techniques based on neuroprotection, anti-inflammatory, nerve stimulation, and anti-stress activation. The molecular genetic alterations in the brain following cerebral ischemia, as well as the mechanism of polypeptide medicines, are unknown. This restricts the use of neuroprotective peptides and makes developing new and more effective medicines to target brain function challenging. Transcriptome analysis is a potential approach for investigating the molecular mechanisms of cerebral ischemia damage and the neuroprotective effects of peptide medications. In addition to investigating the role of mRNA in protein production, researchers must also investigate the role of regulatory RNA in ischemia when developing new neuroprotective methods. MiRNAs and circRNAs, which are mostly expressed in the brain, are the most intriguing. CircRNA may bind with miRNAs and decrease their activity, preventing miRNA-mediated mRNA from being produced. Understanding the circRNA/miRNA/mRNA system is critical for insight into the process of brain damage and healing. Ischemiainduced gene activity changes and peptide-mediated transcriptome spectrum alterations in experimental ischemia were reviewed by Filipenkov et al. [49], who also explained the underlying concept of peptide regulation in ischemia-induced damage. He et al. [50] successfully established the MCAO rat model for the first time and injected 10 mg/kg fluoxetine hydrochloride intraperitoneally for 14 days to study the function of fluoxetine in cerebral IS and the identification of fluoxetine mediated circRNA-miRNA-mRNA axis. After that, triphenyl ammonium tetrachloride staining was used to determine the location of the cerebral infarction. Their findings indicated that fluoxetine might ameliorate brain damage following IS and that the circMap2k1/miR-135b-5p/Pidd1 axis may be implicated.

Summary and prospect
One in every six people worldwide will have a stroke. However, the underlying process is still unknown, and current therapeutic options are limited. CircRNA is a novel form of endogenous non-coding RNA with a covalently closed circular structure. CircRNA, a member of the non-coding RNA family, was originally thought to be incapable of encoding due to a lack of 5'caps and 3'polya tails. Recent research has shown that circRNAs may be translated in a cap-independent manner by acting as the sequence of ribosomal entry sites (IRES) inside circRNAs to facilitate the direct binding of initiation factors or ribosomes to translatable circRNA [51]. CircRNAs have been discovered to be significantly expressed in the central nervous system, indicating that they are nerve specific. Many studies have found that ncRNA expression changes considerably following the onset of AIS. CircRNA is strongly connected to stroke severity and inflammatory response and plays a key role in stroke diagnosis, prognosis, and therapy. Indeed, the molecular and cellular processes that occur following an IS are complicated and staggered. Further studies are required on the use of circRNAs as biomarkers for stroke diagnosis and prognosis. With the development of new sequencing technology and bioinformatics methods, many circRNAs have been identified in different organisms and tissues. These findings have expanded people's understanding of the classification and function of RNAs. The relationship and specific mechanism between CircRNAs and stroke occurrence, development, and prognosis will be gradually clarified. Moreover, the basic principle and molecular mechanism of peptide regulation in ischemia-induced injury will need to be further explored to enable the early detection and intervention of stroke.